The detection of small differences in nucleic acid content is an important task within the field of molecular diagnostics. However, this detection is difficult with current methods.
1 in 20 babies are born with a genetic disorder. Down syndrome is the most common chromosomal disorder affecting about 1 in 750 births (Table 1). The incidence of Down syndrome is increasing with the increasing average age at which women are bearing children.
TABLE 1Incidence and inheritance of fetal aneuploidy.DisorderIncidenceInheritanceDown syndrome1 in 750 birthsTrisomy 21Edward syndrome1 in 3,000 birthsTrisomy 18Patau syndrome1 in 5,000 birthsTrisomy 13Klinefelter syndrome1 in 1,000 births47, XXYTurner syndrome1 in 3,000 births45, XOXYY syndrome1 in 1,000 births47, XYYTriple-X syndrome1 in 1,000 births47, XXX
Down syndrome is caused by trisomy 21—an occurrence of three instead of the normal two copies of chromosome 21. Down patients suffer from mental retardation, heart defects, premature death, and anatomical deformities; most require a lifetime of care. They pose an immense emotional, physical and financial strain on the families and society. Many women therefore want a choice about bringing a child with Down's syndrome into the world or to prepare emotionally for the birth.
Down syndrome is an example of a disease in which early detection is desirable. The tests used today are amniocentesis, chronic villus sampling (CVS), and maternal serum and ultrasound screens.
Amniocentesis is an invasive test requiring an ultrasound-guided needle biopsy of the amniotic fluid surrounding the fetus, through the mother's abdomen. Fetal cells from the amniotic fluid are cultured, and the chromosomes are visualized by fluorescent in-situ hybridization (FISH). Results take 2-4 weeks. Amniocentesis is only recommended between 15 and 18 weeks of pregnancy. It carries a 1% chance of miscarriage and a slight increase in risk of limb disorders. Amniocentesis is estimated to have a sensitivity of 99.3% and a specificity of 99.9%.
Serum screens for Down syndrome are non-invasive tests that measure the level of particalur serum markers. Markers include alpha-fetoprotein (AFP), human chronic gonadotropin (hCG), unconjugated estriol, inhibin A, and PAPP-A. Most markers are tested between the 16th and 18th week of pregnancy, and their combinations have less than 75% sensitivity at a 95% specificity.
In 1979 it was found that maternal blood contains fetal red blood cells (fRBC). In 1997, free fetal DNA was also found in maternal blood serum and plasma (U.S. Pat. No. 5,641,268). These fetal cell and DNA, however, are diluted by significant amounts of maternal cells and DNA (Lo et al., J. Hum Genet. 1998 62, 768-75), complicating the detection of fetal genetic abnormalities.
Cystic fibrosis is a recessive genetic disease. Approximately 30,000 children and young adults suffer from the disease, and one in 31 adults carry a copy of the cystic fibrosis gene in the U.S. This means that one in about 3800 births suffer from cystic fibrosis, which increases to one in every 2,500 births for Caucasians.
Dominant diseases can be detected for a fetus in early pregnancy given a fetal DNA-containing sample, by looking for the presence or absence of the mutant dominant gene. Detecting recessive genetic diseases, such as cystic fibrosis, however, has been difficult because it is not enough to determine if a particular allele is present or not. Rather, it is necessary to determine if the fetus carries 1 or 2 copies of the mutant gene.